Evaluation of Anlotinib Metabolic Stability in HLMs Matrix Applying Ultra‐Fast UPLC–MS/MS Approach: Greenness Assessment With In Silico Testing for Structural Alarms Related to the Metabolic Lability and DEREK Toxicity

Anlotinib (Focus V) is approved by the China Centre for Drug Evaluation in 2018 as a third‐line treatment for patients with certain advanced cancers. This study aimed to develop a specific, fast, reliable, and sustainable ultraperformance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) approach for estimating anlotinib (ANB) in the human liver microsomes (HLMs) matrix, with the application for the assessment of ANB metabolic stability. The validation processes for the UPLC–MS/MS system complied with US Food and Drug Administration regulations following bioanalytical method validation. The established approach was validated using the HLMs matrix over the range of 1.0 to 4000 ng mL −1 within 1 min. The precision (%RSD) and accuracy (%E) rates for intra‐day and inter‐day measurements ranged from −6.33% to 4.91% and −6.67% to 5.26%, respectively. The StarDrop software package involved P450 and Deductive Estimation of Risk from Existing Knowledge modules that were applied for assessing metabolic lability and characterizing ANB structural alarms, respectively. The in vitro half‐life ( t ½ ) was computed at 44.72 min, while the clearance intrinsic (Cl int ) of ANB was calculated at 18.13 mL min −1 kg −1 . In silico evaluations suggest that minor structural alterations to the methyl group (62%), amine group (37%), and methoxy group (1%) in drug design may increase the ANB metabolic stability.